Assessing and Treating Anxiety in Individuals with Autism

Anxiety is a cluster of responses that can involve both operant and respondent behavior, which can be both public and/or private in nature, and occurs when an upcoming aversive stimulus is signaled. Despite the reported high comorbidity of autism and anxiety, there has been very limited research on how to directly assess and treat anxiety, especially with individuals who have limited communication skills. In Study 1, anxiety was assessed in five individuals with autism, ranging in age from 10 to 19 years old. Anxiety was assessed by measuring behavior during (1) a baseline (with no putative anxiety-provoking stimuli present), (2) signals for an upcoming aversive event, and (3) exposure to that aversive event. Anxiety presented in several different ways, as both conditioned activation and suppression, and both with and without problem behavior during the aversive event. In Study 2, individualized treatments involving differential reinforcement of alternative responses and stimulus fading were used to successfully reduce anxious responding in all four participants who displayed anxiety. These studies demonstrated a potentially useful means of assessing anxiety in individuals with autism which may not only help to measure anxious behavior and identify anxiety-provoking events, but may also lead to effective treatment.

Neuropeptidergic regulation of Compulsive Ethanol Seeking in C. elegans

An improved understanding of the molecular basis of alcohol seeking despite the catastrophic consequences of alcohol abuse is likely to enrich our treatments for Alcohol Use Disorders (AUD) and comorbidities. The compulsive seeking is characterized by an imbalance between the superior drive to substance and disruption in control of substance use. To model the development of compulsive engagement of alcohol seeking, we exploit two distinct behavioral programs of C. elegans in conflict, ethanol preference and avoidance of aversive stimulus, simultaneously. We demonstrate that C. elegans exhibited the recapitulation of the pivotal features of compulsive alcohol seeking in mammals, which are repeated attempts, endurance, and finally aversion-resistant ethanol seeking. We find that the neuropeptide signaling via SEB-3, CRF receptor-like GPCR, facilitates the development of ethanol preference and compels animals to seek ethanol compulsively. Furthermore, our functional genomic approach and behavioral elucidation suggest the interaction between neuropeptidergic signaling, SEB-3 and TKR-1, Neurokinin receptor orthologue, to progress compulsive ethanol seeking behavior.

Aversive View Memory and Navigational Risk Sensitivity in the Desert Ant, Cataglyphis Velox

Many ant species are able to establish routes between goal locations by learning views of the surrounding visual panorama. Route formation models have, until recently, focused on the use of attractive view memories, which experienced foragers orient towards to return to the nest or known food sites. However, aversive views have recently been uncovered as a key component of route learning. Here, Cataglyphis velox rapidly learned aversive views, when associated with a negative outcome, a period of captivity in brush, triggering an increase in hesitation behavior. These memories were based on the accumulation of experiences over multiple trips with each new experience regulating foragers hesitancy. Foragers were also sensitive to captivity time differences, suggesting they possess some mechanism to quantify duration. Finally, we characterized foragers perception of risky (variable) versus stable aversive outcomes by associating two sites along the homeward route with two distinct schedules, a fixed duration of captivity or a variable captivity duration, with the same mean time over training. Foragers exhibited significantly less hesitation to the risky outcome compared to the fixed, indicating they perceived risky outcomes as less severe. Results align with a logarithmic relationship between captivity duration and hesitation response, suggesting that foragers perception of the aversive stimulus is a logarithm of its actual value. We conclude by characterizing how view memory and risk perception can be executed within the mushroom bodies neural circuitry.

Associative learning and memory through metamorphosis in Grapholita molesta (Busck) (Lepidoptera: Tortricidae)

Learning of chemical stimuli by insects can occur during the larval or adult life stage, resulting in changes in the imago chemotaxic behaviour. There is little information on learning in Tortricidae, and associative learning through metamorphosis is unknown in this group. Here, we evaluate the influence of olfactory aversive learning in Grapholita molesta (Busck) (Lepidoptera: Tortricidae) during the immature stage and determine if memory persists after metamorphosis. Larvae (10–12 days old) were conditioned to associate the odour of ethyl acetate with pulses of aversive electric shock. Insects were exposed to air, to the ethyl acetate odour, and to shock, in isolation or combination. After conditioning, both larvae and adults were tested in a two-choice olfactometer. Larvae exposed only to air or ethyl acetate increased legibility. Larvae trained with ethyl acetate and shock simultaneously exhibited significant avoidance to ethyl acetate. Avoidance was still present for at least 72 hours after metamorphosis. Thus, G. molesta has the ability to associate an odour to an aversive stimulus precociously, and this association is maintained through metamorphosis and persists into adulthood.

Incorporating inter-individual variability in experimental design improves the quality of results of animal experiments

Inter-individual variability in quantitative traits is believed to potentially inflate the quality of results in animal experimentation. Yet, to our knowledge this effect has not been empirically tested. Here we test whether inter-individual variability in emotional response within mouse inbred strains affects the outcome of a pharmacological experiment. Three mouse inbred strains (BALB/c, C57BL/6 and 129S2) were behaviorally characterized through repeated exposure to a mild aversive stimulus (modified Hole Board, five consecutive trials). A multivariate clustering procedure yielded two multidimensional response types which were displayed by individuals of all three strains. We show that systematic incorporation of these individual response types in the design of a pharmacological experiment produces different results from an experimental pool in which this variation was not accounted for. To our knowledge, this is the first study that empirically confirms that inter-individual variability affects the interpretation of behavioral phenotypes and may obscure experimental results in a pharmacological experiment.

Preference and Motivation Tests for Body Tactile Stimulation in Fish

We tested whether territorial fish (Nile tilapia) perceive body tactile stimulation as a positive or negative resource. Individual male fish were placed for eight days in an aquarium containing a rectangular PVC frame, which was filled with vertical plastic sticks sided with silicone bristles in the middle of the tank. Fish passing this device received a tactile stimulus. The fish then underwent a preference test by choosing between areas half-with and half-without tactile bristles. Then, fish were submitted to a motivation test where they had to pass an aversive stimulus (bright light) to access the device. Fish were, then, paired to settle social rank, which occurs by way of fights (social stressor), and were assigned again to preference and motivation tests. A group without social stress was used as a control. Contrary to our expectations, fish preferred the area without tactile bristles, although subordinate fish reached tactile stimulation more than the dominant one. Social stress did not affect the preference and motivation, suggesting that fish do not perceive tactile stimulation as a stressor reliever. However, as fish did not avoid the stimulation, reached the device spontaneously, and faced an aversive stimulus to access it, we conclude that tactile stimulation is not a negative condition and, therefore, can be used in further studies regarding fish welfare.

Aversive emotion rapidly activates orexin neurons and increases heart rate in freely moving mice

The perifornical area of the hypothalamus has been known as the center for the defense response, or fight-or-flight response, which is characterized by a concomitant rise in arterial blood pressure, heart rate, and respiratory frequency. It is well established that orexin neurons, which are located in this region, play a critical role in this response. In this study, we further examined this role by recording orexin neuronal activity and heart rate in freely moving mice using an original dual-channel fiber photometry system in vivo. Analysis of orexin neuron activity in relation to autonomic responses to aversive stimuli revealed a rapid increase in neuronal activity just prior to changes in heart rate. In addition, we examined whether orexin neurons would be activated by a conditioned neutral sound that was previously associated with aversive stimulus. We show that the memory of the aversive stimulus activated orexin neurons and increased heart rate. Our data suggest that orexin neurons are a key component linking aversive emotions to autonomic defense response. Our data also suggest that targeting orexin neurons may enable treatment of psychiatric disorders associated with chronic stress and traumatic memories.

Enhanced processing of aversive stimuli on embodied artificial limbs by the human amygdala

Body perception has been extensively investigated, with one particular focus being the integration of vision and touch within a neuronal body representation. Previous studies have implicated a distributed network comprising the extrastriate body area (EBA), posterior parietal cortex (PPC) and ventral premotor cortex (PMv) during illusory self-attribution of a rubber hand. Here, we set up a fMRI paradigm in virtual reality (VR) to study whether and how threatening (artificial) body parts affects their self-attribution. Participants (N=30) saw a spider (aversive stimulus) or a toy-car (neutral stimulus) moving along a 3D-rendered virtual forearm positioned like their real forearm, while tactile stimulation was applied on the real arm in the same (congruent) or opposite (incongruent) direction. We found that the PPC was more activated during congruent stimulation; higher visual areas and the anterior insula (aIns) showed increased activation during aversive stimulus presentation; and the amygdala and pregenual anterior cingulate cortex (ACC) were more strongly activated for aversive stimuli when there was stronger multisensory integration of body-related information (interaction of aversiveness and congruency). Together, these findings suggest an enhanced processing of aversive stimuli within the amygdala when they represent a threat to body integrity.

A model of naturalistic decision making in preference tests

Decisions as to whether to continue with an ongoing activity or to switch to an alternative are a constant in an animal’s natural world, and in particular underlie foraging behavior and performance in food preference tests. Stimuli experienced by the animal both impact the choice and are themselves impacted by the choice, in a dynamic back and forth. Here, we present model neural circuits, based on spiking neurons, in which the choice to switch away from ongoing behavior instantiates this back and forth, arising as a state transition in neural activity. We analyze two classes of circuit, which differ in whether state transitions result from a loss of hedonic input from the stimulus (an “entice to stay” model) or from aversive stimulus input (a “repel to leave” model). In both classes of model, we find that the mean time spent sampling a stimulus decreases with increasing value of the alternative stimulus, a fact that we linked to the inclusion of depressing synapses in our model. The competitive interaction is much greater in “entice to stay” model networks, which has qualitative features of the marginal value theorem, and thereby provides a framework for optimal foraging behavior. We offer suggestions as to how our models could be discriminatively tested through the analysis of electrophysiological and behavioral data.Author summaryMany decisions are of the ilk of whether to continue sampling a stimulus or to switch to an alternative, a key feature of foraging behavior. We produce two classes of model for such stay-switch decisions, which differ in how decisions to switch stimuli can arise. In an “entice-to-stay” model, a reduction in the necessary positive stimulus input causes switching decisions. In a “repel-to-leave” model, a rise in aversive stimulus input produces a switch decision. We find that in tasks where the sampling of one stimulus follows another, adaptive biological processes arising from a highly hedonic stimulus can reduce the time spent at the following stimulus, by up to ten-fold in the “entice-to-stay” models. Along with potentially observable behavioral differences that could distinguish the classes of networks, we also found signatures in neural activity, such as oscillation of neural firing rates and a rapid change in rates preceding the time of choice to leave a stimulus. In summary, our model findings lead to testable predictions and suggest a neural circuit-based framework for explaining foraging choices.

Validation of a combined approach-avoidance and conditioned stimulus aversion paradigm for evaluating aversion in chickens

Understanding animals’ aversion is important to improving their welfare. Aversion is often assessed using an approach-avoidance (AA) test in which animals have to forfeit a reward if they want to avoid an event or environment presented in the same place. However, sometimes the event/environment suspected to be aversive may physically impair the animal’s ability to withdraw from that place (i.e. its ability to express aversion), leading to incorrect interpretations. Combining AA with a Conditioned-Stimulus that predicts the event/environment may overcome this problem by allowing animals to demonstrate aversion without exposure to the stimulus. We aimed to validate this paradigm for testing aversion in chickens. Seven Hyline-Brown chickens were trained to obtain a food reward from a coloured bowl located in the test chamber (TC) of a two-chambered box; the reward was presented in a green bowl with an inactivated air canister or a red bowl with the canister activated to deliver an air puff. Two 5-minute tests were conducted, one with each bowl colour and both with the canister inactivated. All chickens entered TC with the green bowl. With the red bowl, two chickens entered on their first attempt, one fully entered after a partial entry (3/7 fully entered), two made only partial entries and two made no attempts to enter. Chickens spent less time in the TC with the red bowl (median 31s, IQR 7–252) compared to the green bowl (293s, IQR 290–294; p = 0.008). The higher ratio of partial to full entries, failure to enter the TC and less time spent in TC reflected chickens’ aversion to the air puff, signalled by the red bowl. The paradigm allowed chickens to demonstrate aversion without exposure to the aversive stimulus during testing.